How do centrifuges work? - Explain that Stuff Tweet by Chris Woodford . Last updated: August 31, 2016. If you need to wash and dry a pair of jeans in a hurry, you'll be awfully glad you have a centrifuge. That's what your clothes washer becomes when it spins wet laundry at high speed to remove the water . A centrifuge is simply a machine that spins around to make a large and useful force. Small centrifuges are used in scientific laboratories (for example, to separate blood products). You can find much bigger ones in aerospace-labs, where they're used for testing astronauts, pilots, and their equipment to absolute breaking point. Let's take a closer look! Photo: The medium-sized space-station centrifuge used by NASA. This one has a radius of 1.8m (6 ft). Photo by courtesy of NASA Ames Research Center and Wikimedia Commons . What is a centrifuge? Photo: A clothes washer drum is a type of centrifuge. During the wash cycle, the paddles agitate the clothes in the soapy water. When it comes to the spin, holes in the drum let the water out. Hold something heavy in one hand and whirl your arm around your head.

Feel a force that seems to be pulling your shoulder out of its socket? That's the principle of the centrifuge at work—and you can look at it from two different angles. In popular books and magazines, people talk about something called centrifugal force: the force that seems to make things shoot outward when they go round in a circle. So, when a bus goes around a bend at high speed, you'll read that it's centrifugal force trying to tip the thing over. When your clothes are spinning in the drum, it's centrifugal force that throws the water out through the little holes so your washing ends up much drier. Or is it? Centrifugal force or centripetal force? Science teachers will tell you this is wrong: there is actually no such thing as centrifugal force. We can understand what's really happening by considering Isaac Newton's famous laws of motion . When a car begins to enter a bend, its natural tendency is to keep going in a straight line—and it will do so unless a force acts on it. When it follows the bend, it does so because there's a force (called centripetal force) constantly tugging it inward from its straight line course.

What we see as the centrifugal force is really the car's tendency to go straight, if left to its own devices. Anytime you hear people talking about centrifugal force, you can quietly correct them (in your own mind, to be polite!) and translate what they're saying into centripetal force. So "centrifugal force gets your washing dry because it makes the water fly out" becomes "Centripetal force between your clothes and the inside of the drum pushes them around in a circle. There's nothing to give the water the same kind of push because it can slip straight through the drum holes. The clothes experience centripetal force, the water doesn't. The clothes go round in a circle, the water goes in a straight line—straight through the holes. And that's what gets your washing dry." Confused? Don't be! It's this simple: if something is moving in a circle, there must be a force acting on it somewhere to make it turn, otherwise it would go in a straight line. So look at the situation carefully and figure out where the inward pushing or pulling force is coming from. That's the centripetal force.

If some part of the object is flying outward, that's not because there's centrifugal force: it's because there's no centripetal force to make it go in a circle. Comparing centrifugal and centripetal force A car's going around a bend in the road. How do we explain what's happening? Centrifugal force: Left to its own devices, the car would go in a circle (blue) but centrifugal force (purple) is constantly trying to push it outward. (The trouble with this is that it doesn't explain why the car is going in a circle.) Centripetal force: Left to its own devices, the car would go in a straight line (orange), but centripetal force pulls it inward (green), so it actually goes round in a circle (red). You can also see now where these two funny words come from. Centrifugal (pronounced sen-tree-few-gul or sen-trif-ugul is related to the word "fugue", from the latin word for flight, so we're talking about something trying to get away from the center.

Centripetal (pronounced sen-trip-itul or sen-tree-pee-tul) comes from the Latin "petere", meaning seek, so we're describing a force that makes things seek the center: Centrifugal = "center fleeing" Centripetal = "center seeking" How big is centripetal force? Why is there always a centripetal force when things move in circles? Think back to the laws of motion . If something is following a curved path, its velocity is changing all the time because its direction is changing all the time. If the velocity is changing, it's accelerating (even if its speed is constant). If it's accelerating, a force must be acting—sometimes a very big force. All these statements follow directly from the laws of motion. Photo: Now that's what I call a centrifuge! You can see how big it is from the little man standing in the bottom, center, wearing a red safety hat. This is the launch-phase simulator that NASA uses to see if pieces of spacecraft (and even whole satellites) can withstand extreme forces.

The huge spinning arms are powered by two 1250-horsepower motors and produce forces up to 30g (30 times the force of gravity or 15 times the force you feel on a typical roller coaster ). Photo by courtesy of NASA Goddard Space Flight Center and NASA on the Commons . Spin things at high speeds and you build up big forces very quickly. You'll have seen that very clearly if you've ever washed something like a heavy pair of curtains in a clothes washing machine and they've bunched up on one side of the drum. When it comes to the final spin, the machine will be banging and clattering about and doing its level best to escape from your kitchen (probably damaging itself in the process). When things spin at high speed, the forces involved can be huge. That's why it's best to balance your washing machine with a mixed load (never, ever wash one item by itself) and why you need to balance laboratory centrifuges very carefully as well. How big are centripetal forces, exactly?

The laws of physics tell us that the centripetal force needed to make an object go round in a circle is given by this little equation: F =(mv2)/r Here, m is the mass of the object, v is the velocity, and r is the radius of the circle. So the bigger the mass of the object and the faster it goes, the more force is needed to keep it turning. Let's try out some real numbers. Say you have a 1000kg car going round a bend of 50m radius at 40mph (roughly 20m/s). The force between the tires and the road is 1000 x 20 x 20 / 50 = 8000 newtons or roughly 10 times a typical person's weight. Where does the centripetal force come from when a car goes round a bend? There's only one place it can come from: the friction between the tires and the road. Ten times a person's weight is quite a lot of force for four little rubber tires to provide, especially when you consider that only a tiny patch of each tire is ever touching the road surface.

Now make the bend twice as tight and you'll see the force is doubled to 16,000 newtons (because r is halved). If the car doubles its speed (v is doubled), the force is quadrupled to 32,000 newtons (40 times a person's weight—or a force equal to 10 people weighing on each tire). That's why you're much more likely to skid at high speeds: friction between your car tires and the road can't provide enough centripetal force to keep you going in a circle, so you whiz off at a tangent: you bow to your natural tendency and keep going in a straight line. What are centrifuges used for? Now forget all about the spinning: if you need a big force quite quickly, a centrifuge is a really handy way to generate one. Because you're spinning on the spot, you don't need lots of space. That makes centrifuges perfect for use in scientific laboratories. One of the most common uses for centrifuges is in separating mixtures of things. A washing machine is a mixture of clothes and water and the spinning drum separates those very efficiently.

Laboratory centrifuges are used to separate things like blood, which consists of red blood cells suspended in plasma (a yellowish fluid). Put some blood in a test-tube and spin it at high speed and you separate out these two components very quickly, with the plasma at the top of the tube and the red blood cells at the bottom. (They travel to the bottom because they're heavier, so need more centripetal force to push them round in a circle. The force comes from the bottom of the tube pushing inward against the blood cells clumped there.) You might have seen astronauts and pilots whizzing round in centrifuges too. There's no special reason why they spin round in a circle; that's pretty much irrelevant. What's being tested is their ability to withstand very high forces—and the easiest way to experience that is at the end of a centrifuge beam. You might feel a bit dizzy at the end but, hey ho, this is rocket science after all! Photo: Students try out a bicycle-powered centrifuge at the Museum of Science and Industry (MOSI) in Tampa, Florida. Note how three people sit at equal angles (sixty degrees apart) to make the centrifuge balance properly as it spins.

Of course, if one person is much heavier than another, there's going to be a problem! Photo courtesy of NASA Kennedy Space Center (NASA-KSC) . How does a laboratory centrifuge work? If you've studied biology or chemistry in school, you've probably used a small, laboratory centrifuge like this one. Although it's a fairly simple bit of equipment, it's cleverly and sturdily designed to survive the extremely high forces involved in spinning samples at speeds of 20,000 rpm or more. Looking inside, from the front, we can see that the main parts are a powerful motor (orange, center); a spindle and flexible collar (blue); and multiple pairs of diametrically balanced pockets into which standard-sized test tubes can be inserted (colored red and green). You load up the machine and securely close the lid, which usually has a lock on it to prevent the centrifuge from being fired up when it's open. Then you switch on by turning a rotary dial on the outside of the case (not shown), which lets you adjust the speed up or down as necessary.

Balance is absolutely critical in a high-speed centrifuge: if one of the pairs of pockets is empty and the other full, the difference in weight will make the machine vibrate dangerously, possibly smashing the sample tubes inside. But balancing test tubes exactly can be very difficult, so machines like this one have flexible rubber collars on their spindles that allow the loaded pockets to rotate about their true center of gravity, thus minimizing any unwanted vibration. Artwork: Inside a typical high-speed laboratory centrifuge. From US Patent 2,699,289: High-speed centrifuge by Robert R. Allen, Custom Scientific Instruments Inc., patented January 11, 1955, courtesy of US Patent and Trademark Office. And this is what it looks like in reality... Photo: Left: Loading up a laboratory centrifuge with samples in test tubes. Note that you can do lots of test tubes at the same time, but you have to make sure the machine is balanced with matching numbers of tubes on each side of the spinning arms. Photo by Scott H. Spitzer courtesy of US Air Force. Right: This is what samples look like when they've been centrifuged.

The lighter components (almost transparent here and quite hard to make out) are at the top of the tube, while the heavier ones (darker here) are at the bottom. Photo by Jim Yost Photography courtesy of US DOE/NREL (Department of Energy/National Renewable Energy Laboratory). centrifuge - Dictionary Definition : Vocabulary.com Random Word centrifuge That ride at the fair that spins you around until you stick to the walls? The device in your washing machine that spins your clothes around quickly to get the water out of them? Those are centrifuges. Centrifuges are used mostly in science. In this application, centrifugal force — the force from spinning that moves things away from the center — separates liquids that have different weights. For example, a centrifuge is used to separate blood cells from plasma cells. When the blood is spun in the centrifuge, the heavier plasma cells separate from the lighter blood cells, and can be collected for other uses. Introduction to the Centrifuge: the Pellet and the Supernatant | Protocol You have full access to this article through University of Washington. The centrifuge is an instrument used in nearly every biomedical research lab across the globe.

Centrifugation is a process by which a centrifuge is used to separate components of a complex mixture. By spinning laboratory samples at very high speeds, the components of a given mixture are subjected to centrifugal force, which causes more dense particles to migrate away from the axis of rotation and lighter ones to move toward it. These particles can sediment at the bottom of the tube into what’s known as a pellet, and this isolated specimen, or the remaining solution, the supernatant, can be further processed or analyzed. The principle component of a centrifuge is the rotor, which is the moving part that spins at high speeds. Rotors can be fixed in position, or a centrifuge can use multiple rotors fixed atop part called the spindle. Usually, the centrifuge rotor will have a lid that is screwed down tightly to prevent samples from flying out. Many centrifuges will have a refrigeration unit that allows the internal temperature to be controlled during the spin. They also have knobs or buttons for inputting the parameters for each run, which can include the duration of the spin, the temperature, and the magnitude of the spin in terms of speed or relative force. Spin speed is measured as RPM, or revolutions per minute.

RPM is tricky value, because it’s not the speed of the centrifuge that causes particles to separate from a mixture, but the force acting on these particles. The force acting on a particle is related to the radius of the centrifuge rotor, and since different centrifuges have different rotor sizes, different forces can be applied at the same speed, or RPM. Centrifugation strength can also be quantified as Relative centrifugal force, or RCF. RCF is generally presented as a multiple of earth’s gravitational acceleration. RCF is expressed as the product of the radius of the rotor and the square of the angular velocity divided by earth’s gravitational acceleration. RCF can be related to RPM by the following equation, where r stands for the radius of the centrifuge measured in centimeters. Using this equation can mean the difference between a failed or successful experiment, but you don’t have to apply this calculation for every new procedure. Centrifuges often come with nomograms that can help convert RCF to RPM quite easily. Use a ruler to connect the radius of the centrifuge and a given RPM value, in order to obtain RCF. To begin spinning your samples consider temperature.

If you are using a refrigerated centrifuge, then you will want the machines internal temperature to reach the desired value before starting the spin, or you can find other ways to cool the rotor. Immediately before a spin ensure all of the caps on your tubes are tightened and secure. When you loading your tubes, ensure that each sample is counterbalanced with another sample directly across from it. If you’ve only got one tube, then make another tube that can act as a counterweight. If you’ve got three tubes, you can arrange them in a triangle so they are equidistant from each other. Balancing weights in a centrifuge is critical. Centrifuge rotors reach high speeds and have a lot of kinetic energy. If improperly balanced, the entire centrifuge unit can be propelled from its resting place and do serious damage. Once you confirm that the rotor and lid are secure, start the centrifuge and hang around until it’s reached the desired speed. If you notice a problem call and experienced member of the lab to come help you. When your centrifugation is complete, you should be able to see your biological specimen at the bottom of the tube in a pellet, which has separated from the rest of the solution, or supernatant.

The supernatant can be removed by either decanting it – a fancy name for pouring it off, or it can be aspirated – a fancy term for using suction to remove it. The purified specimen can then be returned to a solution via a process called, resuspending. Repetitions of centrifuging, or spinning cells, followed by aspirating cells, and resuspending in buffer, is often referred to as, washing cells. Now that you’ve seen some basics of centrifugation, its time to have a look at some of the types of centrifuges out there and the procedures you can carry out with them. Fixed angle rotor centrifuges are probably the most common type of this instrument you’ll encounter in the lab. Many table top centrifuges fit in this category. These centrifuges, in which tubes sit in a fixed and angled position, are used in differential centrifugation protocols. In these protocols, a series of centrifugations at different speeds can be used to purify biological specimens like animal cells. Typically, these protocols involve several cell washing steps. In contrast to fixed angle rotors, swing rotors have flexible tube holders allow samples to rotate outward.

These rotors are beneficial in applications like density gradient centrifugation, where biological samples migrate to distinct layers of gradient media. This type of centrifugation is useful for quickly isolating one cell type from another, or for isolating individual organelles. Lastly, the ultracentrifuge is the big brother of all the centrifuges you’ll find in the lab. It can spin in excess of 70,000 rpm, which makes it well suited for the isolation of small particles, like DNA or viruses. Because of the high speeds of this centrifuge extra care should be taken to ensure that the loads are properly balanced and that both the rotor and lid are secure. You’ve just watched JoVE’s introduction to Centrifugation. In this video we reviewed: what a centrifuge is and how it works, how to operate and run a centrifuge, some safety precautions, and different applications of your centrifugation. Thanks for watching and remember to balance your tubes. Summary The centrifuge is an instrument used in nearly every research lab across the globe. Centrifugation is the process by which a centrifuge is used to separate components of a complex mixture.

By spinning laboratory samples at very high speeds, the components of a given mixture are subjected to centrifugal force, which causes more dense particles to migrate away from the axis of rotation and lighter ones to move toward it. These particles can sediment at the bottom of the tube into what’s known as a pellet, and this isolated specimen, or the remaining solution, the supernatant, can be further processed or analyzed. This video is meant to introduce a student to some of the basic principles of centrifugation, as well as the instrument’s basic operation. For example the speed of centrifugation in rotations per minute, or RPM, is contrasted with relative centrifugal force, or RCF, as a measurement of the magnitude of centrifugation, which is independent of rotor size. In addition to concepts and basic use, safety precautions relating to centrifugation are discussed, as well as the types of centrifuges and centrifuge rotors that exist.

How to Use a Centrifuge: 14 Steps (with Pictures) - wikiHow How to Use a Centrifuge Two Methods: Preparing and Loading Samples Using the Centrifuge Community Q&A A centrifuge is a type of research equipment that spins a liquid suspension at high rotation rates to separate it into distinct layers based on density. [1] Because of these high rotation rates, centrifuges are delicate, can break easily, and can be dangerous when used improperly. Following some simple operating guidelines should prevent any injuries during their use. Steps Preparing and Loading Samples 1 Inspect centrifuge bottles/tubes for cracks before use. [2] Because a centrifuge can spin at such high speeds, a liquid sample can easily become an aerosol if it is not properly contained. Cracked tubes can fracture at high speeds or at the very least, leak sample into the rotor. Discard broken or cracked tubes to prevent them from accidentally being used in the future. 2 Cap tubes with the proper lid. Tubes that have been specified for use with a centrifuge have a proper cap or lid that seals the tube. Using a different lid or covering such as saran wrap or foil can easily lead to spills within the centrifuge.

[3] Making an aerosol of your sample can be hazardous to your health and the environment. In addition, it can lead to loss of precious samples. 3 Clearly label tubes for identification. You may know how you placed your tubes into the centrifuge before they started spinning, but at the end of the spin you will not be able to tell them apart. Make sure you put some sort of label on each tube so you know which sample is which. [4] If you have a balance, be sure you label it as such. It is best to label the tube directly instead of using a sticker. A sticker can fall off during the spin making identification difficult. 4 Wipe the outside of the tube with disinfectant before placing it in the centrifuge. Wiping the tube is particularly important if you are working with a biohazardous material. [5] You want to limit any possible spills or aerosol formation of your sample. The best prevention is to wipe down the sample with a proper disinfectant before the spin begins. 5 Make a counterbalance for the centrifuge tube you want to put in the centrifuge. At high speeds, a centrifuge can easily become unbalanced if there aren’t equal masses opposite each other in the rotor.

Remember to balance the masses of the tubes, not the volumes. Weigh the tube with your sample and record the mass. Add water to a separate empty tube until the mass is the same as your sample. [6] Unbalanced tubes can lead to permanent damage of the centrifuge and can be hazards if the rotor breaks free from the rest of the centrifuge. Balancing is extremely important at higher centrifugation speeds. Many centrifuges will automatically turn off if it senses an unbalanced load, but older ones may not have this feature. If the centrifuge begins to shake or wobble, it is off balance and you should stop it immediately. Method Using the Centrifuge 1 Place the centrifuge on a firm, level surface. Because of the high speeds the centrifuge spins at, it needs to be housed on a solid, flat surface. Find a level table or counter that is secure and can properly support the weight of the centrifuge. If you notice the centrifuge sliding around or the counter underneath sagging, relocate the centrifuge to a more stable place. 2 Choose the proper rotor to use at the speed you need. The rotor is the piece that spins your sample.

Some rotors have arms that you can attach smaller buckets to, while other rotors are one piece that you place the tubes directly into. Rotors are designated for specific tubes and speeds. Use a rotor that fits the tubes of your sample and is within the limit of speed needed. If your samples are in the wrong type of tube, you may need to transfer them to the proper type before spinning. 3 Load the tubes opposite each other in the centrifuge. If you are only spinning one sample, you will need to make a balance to load directly opposite the sample tube. [7] If you are spinning more than two tubes, only the ones directly opposite each other have to be equal in mass. 4 Enter the centrifugation speed. Centrifugation speed is often given in rotations per minute (rpm), or how many times the rotor completes a full rotation in one minute. Speed can also be given in relative centrifugal force (RCF) or the G-force on the rotor. [8] Modern day centrifuges are digital and have settings for both. The speed at which you spin your samples is dependent upon what you are spinning. 5 Keep a safe distance while the centrifuge is running.

Bumping or moving the centrifuge during use can cause an imbalance and lead to injury. [9] Larger centrifuges can spin at very high speeds and are extremely dangerous if they become imbalanced. 6 Turn off the centrifuge if it is wobbling. After starting the centrifuge, stay close until it gets up to full speed in case it needs to be turned off. When a centrifuge is off balance it may shake or wobble. This can damage the equipment and can also lead to injury in severe cases. If you cannot get to the power button, simply unplug the centrifuge. [10] Many centrifuges will automatically turn off if it senses an unbalanced load, but older ones may not have this feature. If the centrifuge begins to shake or wobble, it is off balance and you should stop it immediately. A small amount of vibration is normal, but excessive wobbling is dangerous. If your samples are balanced and the wobble still occurs, stop using the centrifuge and call for maintenance. 7 Open the lid only after the rotor has completely stopped. [11] Many modern centrifuges have a lock on them that will not unlock until the rotor has come to a stop.

Older centrifuges may not have this feature so make sure the rotor is no longer moving before opening the lid. Keep power cords away from the ground to avoid trip hazards that could move the centrifuge. 8 Remove the tubes carefully after the centrifuge has completely stopped spinning. You want to gently remove the tubes so that the separated suspensions do not mix again. You also want to check to see if any of the samples have leaked or tubes have broken. If a spill occurs, clean rotor and buckets immediately. 9 Wipe down rotor and centrifuge after each use. To keep the centrifuge clean and running smoothly, lightly clean the rotor and wipe down the centrifuge at the end of a run. When you are finished, leave the lid open so it can air out and remain dry. Community Q&A Will the imbalance or speed of a centrifuge cause a serum to turn a pink color? wikiHow Contributor The imbalance of a centrifuge would cause test tubes to break. The speed determines how quickly the product would be centrifuged, and may change the appearance/thickness of the outcome. How do I balance the load if there is only one sample test tube? wikiHow Contributor The tube opposite your sample tube should be filled with water of equal mass.

It's very important that the mass is the same, not the volume. If this question (or a similar one) is answered twice in this section, please click here to let us know. Video Tips When balancing your tubes, it's helpful to put a small beaker or a similar object on the balance to hold your tubes upright so you don't have to take them off the balance to add water or other solvents. Keep the lid closed while the centrifuge is spinning. Always wear gloves. Warnings Unbalanced centrifuges may slowly move across the surface it's on. Immediately shut down unbalanced centrifuges to prevent damage to the machine, you, and those around you. Things You'll Need Chemistry for Kids: Separating Mixtures Chemistry for Kids Separating Mixtures Many of the substances we use everyday were actually once part of a mixture. Someone somewhere separated that substance from the mixture so we could use it. It turns out that many compounds and elements aren't found in nature in their pure form, but are found as parts of mixtures. Separating substances from mixtures is an important part of chemistry and modern industry. Some important chemistry terms are used in this section including mixtures , suspensions , and solutions .

You can click on the links to learn more about each of them. Why do we want to separate mixtures? All the way back to Ancient History, industrious humans have separated mixtures in order to obtain the specific substances that they need. One example of this is extracting metal from ore in order to make tools and weapons. We'll discuss some other examples of separation below. Separation Processes The way in which different substances in a mixture are separated is called a process. There are a number of different processes used for separation. Many of them are very complex and involve dangerous chemicals or high temperatures. A lot of important industries in the world today are based on separation processes. Filtration One common method of separation is filtration. Filters are used everywhere. We use them in our houses to filter dust and mites out of the air we breathe. We use them to filter impurities from our water. We even have filters in our bodies such as our kidneys which act as filters to get bad stuff out of our blood. The filtration process is generally used to separate a suspension mixture where small solid particles are suspended in liquid or air. In the case of filtering water, the water is forced through a paper that is made up of a very fine mesh of fibers.

The water that has been run through the filter is called the filtrate. The particles that are removed from the water by the filter are called the residue. Distillation Another common separation process is called distillation. Distillation uses boiling to separate mixtures of liquid solutions. It takes into account that different substances in the mixture will have different boiling points. For example, if you heat salt water the water in the solution will boil before the salt. The water will then evaporate leaving the salt behind. If the steam from the water is collected it will turn back into liquid as it cools. This cooled water will be pure water without any salt. Centrifuge In some cases, there are suspension mixtures where the solid particles are too fine to be separated with a filter. In these cases, sometimes a centrifuge is used. Centrifuges are mechanical devices that spin at very high speeds. These high speeds allow the solid particles in suspensions to settle very quickly. For example, rather than wait for sand to slowly settle to the bottom of water, a centrifuge can cause the sand to settle in a matter of seconds.

Some examples of how centrifuges are used include separating blood into plasma and red cells, separating cream from milk, and separating uranium isotopes for nuclear power plants. The heavier particles move to the outside of the cylinder as the centrifuge spins allowing the mixture to be separated. Other Processes There are many other separation processes such as sublimation, adsorption, crystallization, and chromatography. Sometimes it takes many stages of processes to get to the final result. One example of this is the processing of crude oil. Crude oil uses many levels of fractional distillation to produce a number of different products including gasoline, jet fuel, propane gas, and heating oil. Interesting Facts about Separating Mixtures To separate liquid solutions where the substances have similar boiling points, a more complex version of distillation is used called fractional distillation. Painting uses the separation process of evaporation. The wet paint is a mixture of color pigment and a solvent. When the solvent dries and evaporates, only the color pigment is left. The separation process of winnowing was used in ancient cultures to separate the grain from the chaff. They would throw the mixture into the air and the wind would blow away the lighter chaff, leaving the heavier grain.

High speed centrifuges can spin up to 30,000 times a minute. Many separation processes are occurring constantly in nature. Activities Take a ten question quiz about this page. More Chemistry Subjects On-Line Resources for Biology: Centrifugation Basics Centrifugation Theory Centrifugation is a process used to separate or concentrate materials suspended in a liquid medium. The theoretical basis of this technique is the effect of gravity on particles (including macromolecules) in suspension. Two particles of different masses will settle in a tube at different rates in response to gravity. Centrifugal force (measured as xg, gravity) is used to increase this settling rate in an instrument called a centrifuge. Two common examples of the use of centrifugal force are: (1) When you do the "around the world" trick with a yo-yo, it is centrifugal force that makes the yo-yo body stay at the end of the string as you rotate it; and (2) When you wash clothes in a washing machine, it is centrifugal force generated in the "spin" cycle that forces water out of the fabric to facilitate faster drying.

Centrifuges are devices used in a variety of scientific and technical applications which spin carrier vessels (centrifuge tubes) at high rotation speeds and very high centrifugal force. The centrifugal force (expressed as # gravities or, # xg) generated is proportional to the rotation rate of the rotor (in rpm) and the distance between the rotor center and the centrifuge tube. Therefore, a given centrifuge may use multiple rotor sizes to give flexibilty in choosing centrifugation conditions. Each centrifuge has a special graph, a nomograph, or a table which relates rotation rate (rpm) to centrifugal force (xg) for each size of rotor it accepts. Typically, the material to be "spun" is placed in a centrifuge tube which is then placed in a rotor. The rotor is generally a dense metal which dissipates heat quickly, and is of sufficient mass that it generates momentum, i.e., once its spinning it requires little energy to keep it going. Centrifuges generally work under vacuum and are refrigerated to reduce heating caused by frictional forces as the rotor spins. Rotors are usually stored in refrigeration units to keep them at or near the operating temperature.

Centrifuges come in all shapes and sizes, and the rotors vary, therefore, the universal and transferable unit of centrifugation is centrifugal force in gravities (xg). Different makes of centrifuges use different rotors and each model comes with a table or a graph that relates centrifugal force to rotational speed (rpm) for each rotor (or swing buckets) it can use. In lab write-ups you should ALWAYS report the centrifugal force used (#gravities) and duration of time at that force because centrifugal force is the only transferable unit between different centrifuges. Blood-Separation-and-Centrifugation-52896 Page: 1 | 2 | 3 | 4 | 5 | 6 | 7 | Table of Contents | Download PDF There is a relationship between the physical properties of blood; liquid viscosity, particle density, particle size, fraction of dissolved solids and the physical principles of centrifugation (gravitational force applied) that impact separation. Guidelines are provided to improve the efficiency of blood component separation using differential centrifugation. Introduction There is only so much plasma that can be obtained from a unit of whole blood.

Total plasma volume is determined by the volume of blood collected and the donor percent hematocrit (% Hct). Figure 1 illustrates the theoretical range of plasma that can be recovered from a unit of whole blood. Figure 1 Theoretical Total Plasma Volume Dependent on Donor % Hct and Collection Volume Principles of Blood Separation Blood separation is accomplished by sedimentation and can be defined as the partial separation or concentration of suspended solid particles from a liquid by gravity. The rate of sedimentation is a function of liquid viscosity, particle density, particle size, concentration of the solution (fraction of dissolved solids), and the force of gravity. Sedimentation rates can be calculated for any particulate fluid using Stokes Law of Sedimentation.iThis equation states that at any given “g-force”, the rate of sedimentation of a particle is directly proportional to its size and density and relative to the density of the suspension fluid. To accelerate sedimentation, the effect of gravity is amplified using “centrifugal force” provided by a centrifuge and can be many thousand times the force of gravity. Separation of cellular constituents within blood can be achieved by a process known as differential centrifugation.

In differential centrifugation, acceleration force is adjusted to sediment certain cellular constituents and leave others in suspension. During the process of differential centrifugation of blood, the sample is separated into two phases: a pellet consisting of cellular sediment and a supernatant that may be either cellular or cell-free. iStokes Law of Sedimentation Vg = d2 (Þp - Þ1) / 18μ x G where: Vg = sedimentation velocity, d = particle diameter, Þp = particle density, Þ1= liquid density, G = gravitational acceleration, μ= viscosity of liquid Standardizing Centrifugation Terminology Centrifugation terminology can be confusing. Abbreviations such as RPM, RCF, TCF and ACE™ are used to describe centrifugation conditions. As the discussion progresses on plasma yield optimization, it is important to clearly understand common centrifugation terminology. Standardization of centrifugation terminology will simplify the approach used later in this document to optimize centrifugation and improve plasma recovery. Centrifugation terminology is defined below. Revolutions per Minute (RPM) – is the rotating speed of the rotor arm within the centrifuge during centrifugation.

Example: 5000 RPM. The centrifuge is spinning at 5000 Revolutions per Minute. g – Force or “g” – is a unit of acceleration equal to the force of gravity. Relative Centrifugal Force (RCF) – is the force during centrifugation that moves a particulate away from the center of rotation. It is expressed as multiples of the earth's gravitational field (g). Since RCF includes rpm in the calculation, it changes as the rpm changes during the centrifugation cycle. During acceleration the RCF increases, when the rpm set point is attained, the RCF is constant and during deceleration the RCF decreases. Therefore, the RCF calculated during the centrifugation cycle reflects the force applied at any particular instant in time.

The formula to calculate relative centrifugal force in “g” is as follows: RCF (g) = (rpm)2 x (1.118 x 10-5) x radius of rotor (cm) RCF (g) = (rpm/1000)2 x 28.38 x radius of rotor (inches) Total Centrifugal Force (TCF) – is a calculation for determining the total applied centrifugal force over the complete centrifugation cycle and has units of g•s. Whereas RCF is the force applied at a given time in the centrifugation cycle, TCF is the Total Force Applied over the complete time and uses RCF (g) x time (s). TCF (g•s) =RCF (g) x time (s) Accumulated Centrifugal Effect1 (ACE) – is a calculation for determining the total applied centrifugal force over time and uses speed (RPM) and time (s). This calculation can be programmed on most Sorvall™ centrifuges as the “integrator function” or ∫ω2dt.

While centrifugation protocols are frequently defined as RPM and time, this practice can introduce significant variability from one center to another or one centrifuge to another when trying to reproduce the same process. Expressing the force of centrifugation using RPM alone does not take into consideration radius of the rotor or centrifuge load. The variability of centrifugation can be reduced by standardizing the terms of acceleration to specify the acceleration (RCF: g) or total centrifugal force (TCF: g x s) or accumulated centrifugal effect (ACE) that is to be applied to the sample, rather than specifying revolutions per minute and time. Calculation of RCF is dependent upon the radius of the centrifuge rotor used. The same centrifuge manufacturer with different rotors can produce different acceleration forces (defined as multiples of “g” or the earths’ gravitational force). An example of the difference in RCF for the same RPM settings is shown in Table 1. Table 1 Relative Centrifugation Force Values for Various Centrifuge Rotors Centrifuge Manufacturer 2120 6539 Table 1 shows the impact of using RCF and RPM nomenclature interchangeably.

In this example the assumption that centrifugation of blood components at 5000 RPM is equivalent to an RCF of 5000 x g could subject blood components to elevated g-force and therefore compromise the component quality and the efficiency of component separation. Defining the time of centrifugation is also important in standardizing centrifugation protocols. Time may be defined in minutes of acceleration at a given RCF or it may be incorporated in the final calculation of either total centrifugal force2 or accumulated centrifugal effect. Both TCF and ACE calculations are used to standardize the total effect of centrifugation and use a calculation of either speed and time (ACE) or RCF and time (TCF) to calculate the area under the curve (AUC). An example using area under the curve to calculate TCF for centrifugation is shown in Figure 2.

Figure 2 Calculating Total Centrifugal Force Using Area under the Curve Fraction 13968 1.73 x 106 g.s As shown in Figure 2, using AUC to calculate TCF is as simple as multiplying the RCF x time (s) for each incremental fraction (depicted in the graph as fractions 1-9) and adding the individual fractions that make up the curve. A similar method of using time and RPM to calculate area under the curve and associated ACE settings using the integrator function ∫ω2dt is well described1.The benefits of using the ACE function are to standardize centrifugation by improving run-to-run reproducibility. The setting compensates for differences in acceleration associated with differences in rotor load, voltage fluctuations, loss of instrument calibration, and environmental factors like extreme temperatures2. General Overview of Centrifugation for Blood Banking There are three differential centrifugation processes used to separate whole blood into blood components for transfusion. These are: soft or light spin, hard or heavy spin, and buffy coat spin processing conditions. The soft spin centrifugation profile uses low centrifugal force (slower speed) to allow slower sedimentation of the lighter constituents in whole blood.

The soft spin processing condition produces a plasma component that contains a large fraction of the platelets (platelet-rich plasma) and a softly packed red cell component. The buffy coat layer is located at the interface that separates the platelet-rich plasma from the packed red cells, and contains the majority of the white blood cells as well as some of the larger more dense platelets. The soft spin process is typically used to prepare platelets, packed red cells, and plasma from a whole blood component. Hard spin component processing uses high centrifugal force (faster speed and longer centrifugation time) and produces compacted red cell product and low cellular content, high volume plasma. In this case, the buffy coat layer contains most of the blood leukocytes and platelets. If at this stage the buffy coat layer is separated from the plasma and red cells and is subjected to another round of soft spin centrifugation, a buffy coat platelet product can be produced as is currently performed in Canada and many European countries. In the United States, the AABB recommends centrifugation conditions for the production of blood components as outlined in Table 2. Table 2 AABB Recommended Centrifugation Conditions3for Component Separation

Question: Which instrument can separate two liquids by spinning them at high speed?
Answer: Centrifuge